For successful gene therapy and treatment with novel vaccines, efforts are being made today in order to ensure that biopolymers such as genes and antigenic proteins are introduced into the cytoplasm directly and efficiently. Unlike low-molecular weight drugs, high-molecular weight substances such as genes, antigenic proteins and physiologically active proteins are not only low in membrane permeability, absorption and tissue migration but they also undergo rapid degradation in blood. It has been desired to develop a technology by which those polymeric, physiologically active substances can be introduced into the cytoplasm in a desired and efficient manner without causing damage to the cell.
Liposomes can hold a number of substances and are still biocompatible, so they have drawn researchers' attention as carriers for transporting physiologically active substances. However, with liposomes, the efficiency of introducing the desired substance into a cell is sow low that it can hardly be introduced into the cytoplasm. To deal with this difficulty, various proposals have been made, including modifying the surfaces of liposomes with lectins, antibodies, etc. so that they are positively bound to cell surfaces. In fact, however, liposomes, whether surface-modified or not, are taken up by cells via endocytosis, so they are lysed by an enzyme called lysozyme and the proportion in which the substance of interest is actually transferred into the cytoplasm is at an extremely low level.
It was reported that in order to overcome this difficulty, fusogenic liposomes having the fusogenic capability of Sendai virus were developed as carriers that could be directly introduced into the cytoplasm via the cell membrane. Such fusogenic liposomes can be prepared by forming a complex between a liposome and the coat protein of Sendai virus which mediates fusion to the cell membrane. The prepared fusogenic liposomes have an almost comparable fusogenicity to Sendai virus and it has been reported that by encapsulating genes, proteins or other high-molecular weight substances, the substance of interest can be directly introduced into the cytoplasm with high efficiency but without causing cell injury [“DDS”, Journal of the Japan Society of DDS, Vol. 13, No. 1, January 1998, pp. 21-26 and 27-33].
Even if the fusogenic liposomes are used as carriers for physiologically active substances, the release of the physiologically active substance in cells cannot be controlled, so the substance introduced into the cell is released at a time and its activity (toxicity) is not sustained. Take, for example, the case of introducing a gene into the cytoplasm; the gene is decomposed in the cytoplasm and its expression is not sustained. In the case of a protein having pharmacological activity, the activity is not sustained and it has to be administered by an increased number of times in a larger dose. Therefore, if physiologically active substances are introduced into cells directly and efficiently and if their release is controlled within the cytoplasm, the intended physiological activity can be exhibited efficiently in the cell. It is desired to develop a transport carrier that permits a substance of interest to be introduced into cells and which enables slow release of the substance in the cell.
Desired is the development of a safe and stable transport carrier that allows physiologically active substances, in particular high-molecular weight substances such as proteins and genes, to be introduced into cells efficiently and without damaging the cell membrane and which still can control and adjust the release profile of the introduced substance within the cell.